We are exploring gene regulation of rabbit Kappa immunoglobulin genese by characterizing several genomic clones related to this system. The important regions of the nominal b4 allotype Kappa gene have been sequenced (about 5 Kb including complete J-C locus). Other C genes currently under investigation include a gene from b4 DNA encoding the Kappa2 isotype and a gene apparently encoding the nominal b5 allotype Kappa chain. In addition, we are sequencing a germline genomic V region clone and an enigmatic rearranged gene isolated from a rabbit-mouse hybridoma. Probes from cloned genomic DNA have been used to characterize, by Southern blot analysis, Kappa-related sequences in DNA from rabbits of several b series strains. We are also studying the expression of four human genes whose activity is regulated in B cell development. In collaboration with Dr. Stanley Krosmeyer of NCI, several leukemia and lymphoma lines representing B-lineage cells at different stages of differentiation have been examined for the methylation status of four genes; the methylation analysis is being correlated with the magnitude of gene expression as assessed by Northern blot analysis. These studies represent assessment of baseline parameters which we hope to be able to alter by in vitro manipulation of expression of these genes.